Work continues to understand the genetic regulation of expression of proteins secreted by Bacillus anthracis and Bacillus cereus that are potential virulence factors. Proteins of particular interest are hemolysins, toxins, and proteases. In B. cereus, expression and secretion of these is controlled by a global regulator, PlcR, which is part of a quorum sensing system that responds to peptides generated from a small protein, PapR. Work now focuses on identifying the secreted protease that is responsible for generation of the peptides from PapR. A candidate protease was identified by genomic comparisons and it has been inactivated using the Cre-Lox system developed previously for gene knockouts. This gene KO eliminated the quorum sensing, as reflected in the loss of expression of several secreted factors. [unreadable] In other work, efforts are underway to develop improved tools for controlling and detecting heterologous gene expression. Inducible promoter systems that have been useful in other gram-positive bacteria have been adapted for use in B. anthracis. [unreadable] During this period we continued to promote the use of a unique monoclonal antibody, S9.6, that reacts in a sequence-independent manner with DNA/RNA hybrids. Several labs to which we have supplied this antibody have confirmed that it has great value in development of microarrays, in part because it eliminates the need to label or amplify cellular RNAs, procedures that can introduce bias or variation.